Partitioning oil

Manufactured by 10x Genomics

Partitioning Oil is a specialized oil used in the processing of samples for single-cell analysis with 10x Genomics' technology. It plays a critical role in the encapsulation of individual cells or nuclei within nanoliter-scale droplets, which is a key step in the 10x Genomics workflow.

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Lab products found in correlation

4 protocols using partitioning oil

Optimized Chromium Single Cell ATAC Protocols

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The Chromium Single Cell ATAC v.1.0 E Chip (10x Genomics #2000121) was loaded with 80 μl of 1x Nuclei Buffer (10x Genomics #2000153) into inlet 1, 40 μl of Single Cell ATAC Gel Beads (10x Genomics #2000132) into inlet 2, and 240 μl of Partitioning Oil (10x Genomics #220088) into inlet 3. The Chromium Single Cell ATAC v.1.1 Next GEM H Chip (10x Genomics #20000180) was loaded with 70 μl of 1x Nuclei Buffer (10x Genomics #2000153) into inlet 1, 50 μl of Single Cell ATAC Gel Beads v.1.1 (10x Genomics #2000210) into inlet 2, and 40 μl of Partitioning Oil (10x Genomics #220088) into the outlet, labeled as row 3. Because we omitted Reducing Agent B, the gel beads remained intact throughout the microfluidic run, such that they could be visualized inside the emulsion droplets using a standard light microscope. Our fill rate calculations are based on a total of 1,265 (scATAC v.1.0) or 1,610 (scATAC v.1.1 Next GEM) droplets that were manually counted from microscopic images.

Datlinger P., Rendeiro A.F., Boenke T., Senekowitsch M., Krausgruber T., Barreca D, & Bock C. (2021). Ultra-high-throughput single-cell RNA sequencing and perturbation screening with combinatorial fluidic indexing. Nature methods, 18(6), 635-642.

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Scalable Single-Cell RNA-Seq Library Prep

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Single cell RNA-seq libraries were generated using Single Cell 3′ Reagent Kits v2 (10xGenomics) according to the manufacturer’s protocol. Briefly, the Chromium Single Cell A Chip (10xGenomics, 1000074) were loaded with cells, reagents (10xGenomics, PN-120237), gel beads (10xGenomics, 1000092), and partitioning oil (10xGenomics, 220088). cDNA was generated using the Gel Bead-In-Emusions (GEMs)-RT incubation protocol, purified with DynaBeads MyOne Silane beads, amplified with PCR, and further purified with SPRI-select reagent (Beckman Coulter, B23318). After fragmentation, end-repair, A-tailing and size purification, cDNA was ligated with adaptor, followed by quantification with SPRI-select reagents. Libraries were amplified using PCR with sample indexes from Chromium i7 Multiplex kit (10xGenomics, PN-120262). Library quality control was performed on the Agilent Tapestation with D1000 screen tapes, and library yield was quantified by Qubit DNA high sensitivity assay. Sequencing was performed on an Illumina Nextseq 500 system operated by the Next Generation Sequencing at GENEWIZ using the HiSeq 2 × 150 bp high output sequencing kit.

Ali M., LaCanna R., Lian Z., Huang J., Tan Y., Shao W., Yu X, & Tian Y. (2022). Transcriptional responses to injury of regenerative lung alveolar epithelium. iScience, 25(8), 104843.

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Single-Cell RNA-Seq Library Preparation

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A single cell/nuclei suspension containing 10,000 cells/nuclei was loaded on a Chromium Single Cell B Chip (10x Genomics, catalog no. 1000154) as follows: 75 μL of master mix + nuclei suspension was loaded into the row labeled 1, 40 μL of Chromium Single Cell 3 Gel Beads (10x Genomics, catalog no. PN-1000093) into the row labeled 2 and 280 μL of Partitioning Oil (10x Genomics, catalog no. 2000190) into the row labeled 3. This was followed by GEM generation and barcoding, post GEM-RT cleanup and cDNA amplification. Then 100 ng purified cDNA derived from 12 cycles of cDNA amplification was used for 3 Gene Expression Library Construction by using Chromium Single Cell 3 GEM, Library & Gel Bead Kit v3 (10x Genomics, catalog no. 1000092) according to the manufacturer’s manual (10x Genomics, catalog no. CG000183 Rev C). The barcoded short-read libraries were measured using a Qubit 2.0 with a Qubit dsDNA HS assay kit (Invitrogen, catalog no. Q32854) and the quality of the libraries was assessed on a Fragment analyzer (Agilent) using a high-sensitivity NGS Fragment Kit (1–6000bp) (Agilent, catalog no. DNF-474-0500). Sequencing libraries were loaded on an Illumina NovaSeq6000 with PE 2 × 50 paired-end kits by using the following read length: 28 cycles Read1, 8 cycles i7 index and 91 cycles Read2.

Joglekar A., Hu W., Zhang B., Narykov O., Diekhans M., Balacco J., Ndhlovu L.C., Milner T.A., Fedrigo O., Jarvis E.D., Sheynkman G., Korkin D., Ross M.E, & Tilgner H.U. (2023). Single-cell long-read mRNA isoform regulation is pervasive across mammalian brain regions, cell types, and development. bioRxiv.

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Chromium Next GEM Chip H Single-Cell ATAC-Seq

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On the Chromium Next GEM Chip H (10x Genomics #2000180), unused channels were loaded with 70 μl (row 1), 50 μl (row 2) and 40 μl (row 3) of 50% glycerol solution (Sigma #G5516-100ML). Per sample, we added a mix of 41.9 μl of nuclease-free water, 12 μl of 10x Ampligase Reaction Buffer (Lucigen #A0102K), 2.3 μl of 100 U/μl Ampligase (Lucigen #A0102K), 1.5 μl of Reducing Agent B (10x Genomics #2000087) and 2.3 μl of 100 μM Bridge Oligo (sequence provided in Supplementary Table 1) immediately before the run. Seventy microliters of cell suspension in thermoligation master mix was loaded into the Chromium Next GEM Chip H (row 1), along with 50 μl of Single Cell ATAC Gel Beads v.1.1 (row 2, 10x Genomics #2000210) and 40 μl of Partitioning Oil (row 3, 10x Genomics #2000190), and the chip was run on the Chromium system.

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